The bacteriophage ϕX174 has a single-stranded DNA genome of 5386 bases. During DNA replication, double-stranded forms of the genome are generated. In an effort to create a restriction map of ϕX174, you digest the z-stranded form of the genome with several restriction enzymes and obtain the following results. Draw a map of the ϕX174 genome.

Verified step by step guidance

1. Step 1: Understand the data provided. The genome of bacteriophage ϕX174 is 5386 bases long. The table shows the fragment sizes obtained after digestion with different restriction enzymes: PstI, PsiI, and DraI, both individually and in combinations. Single enzyme digests give the total fragment sizes, while double digests give overlapping fragment sizes that help locate the cut sites relative to each other.
2. Step 2: Start by mapping the single enzyme digests. Since PstI and PsiI each cut the genome once (single fragment of 5386 bases), they likely cut at one site each, linearizing the circular genome. DraI cuts twice, producing two fragments of 4307 and 1079 bases, indicating two cut sites.
3. Step 3: Use the double digest data to determine the relative positions of the cut sites. For example, the PstI + PsiI digest produces fragments of 3078 and 2308 bases. Since PstI and PsiI each cut once, these two fragments sum to 5386 bases, confirming the genome length. This tells us the distance between the PstI and PsiI sites is 2308 bases on one side and 3078 bases on the other.
4. Step 4: Similarly, analyze the PstI + DraI digest fragments (331, 1079, 3976 bases) and PsiI + DraI digest fragments (898, 1079, 3409 bases). These three fragments sum to 5386 bases, confirming the genome length. Use these fragment sizes to position the DraI sites relative to PstI and PsiI by matching overlapping fragment sizes and distances.
5. Step 5: Draw the circular genome map by placing the PstI site at an arbitrary position (e.g., 0), then mark the PsiI site 2308 bases away. Next, place the two DraI sites such that the fragment sizes between PstI, PsiI, and DraI sites correspond to the fragment sizes observed in the double digests. This will give a complete restriction map of the ϕX174 genome.

Key Concepts

Restriction Enzyme Digestion and Fragment Analysis

Restriction enzymes cut DNA at specific sequences, producing fragments of varying lengths. By analyzing the sizes of these fragments after digestion, one can infer the locations of enzyme cut sites on the genome. Comparing single and double digests helps determine the relative positions of these sites.

Restriction Mapping

Restriction mapping is the process of creating a physical map of a DNA molecule based on the pattern of fragments generated by restriction enzyme digestion. By combining data from single and multiple enzyme digests, the order and distance between restriction sites can be deduced, allowing construction of a genome map.

Double-Stranded DNA Replication of Single-Stranded Genomes

Although ϕX174 has a single-stranded DNA genome, it forms double-stranded replicative forms during replication. Restriction enzymes require double-stranded DNA to cut, so digestion experiments are performed on these replicative forms to analyze the genome structure.